Team Finds Weak Spot on Deadly Ebolavirus

Scientists
from The Scripps Research Institute and the US Army’s Medical Research Institute
of Infectious Diseases have isolated and analyzed an antibody that neutralizes Sudan
virus, a major species of ebolavirus and one of the most dangerous human
pathogens.

“We
suspect that we’ve found a key spot for neutralizing ebolaviruses,” said
Scripps Research Associate Professor Erica Ollmann Saphire, who led the study
with US Army virologist John M. Dye.

The
new findings, which were reported November 20, 2011, in an advance online edition
of Nature Structural & Molecular
Biology, show the antibody attaches to Sudan virus in a way that links two
segments of its coat protein, reducing their freedom of movement and severely
hindering the virus’s ability to infect cells. The protein-linking strategy
appears to be the same as that used by a previously discovered neutralizing
antibody against the best-known ebolavirus species, Ebola-Zaire. The new study
suggests that this may be the best way for vaccines and antibody-based
therapies to stop ebolaviruses.

Deadly Outbreaks

Ebolaviruses
first drew the attention of the medical world with simultaneous deadly
outbreaks in 1976 in the nations of Sudan and Zaire (currently known as the
Democratic Republic of the Congo). These two outbreaks were caused by the two
major viruses: Ebola-Sudan and Ebola-Zaire, and early field studies showed that
sera from patients that survived one virus could not help patients infected with
the other. . Both viruses persist in animal hosts–probably bats–and when they
spread to humans, typically cause severe hemorrhagic fevers, killing up to 90
percent of people they sicken. Although not as contagious as influenza or
measles, ebolaviruses can be transmitted in bodily fluids including exhaled
airborne droplets, and scientists who study these viruses are generally
required to use special “Biosafety Level 4” facilities. The US government
regards the ebolaviruses as a potential bioterror threat.

Ebolavirus
researchers hope to develop a vaccine that could be used to protect health
workers and others in the vicinity of ebolavirus outbreaks, as well as an
antibody-based immunotherapy that could help infected people survive. However,
these tasks are complicated by the fact that there are now five recognized
species of ebolavirus: Ebola-Zaire, also known simply as Ebola virus; Taï
Forest virus; Reston virus; Bundibugyo virus; and Sudan virus.

“These
species differ enough from each other that neutralizing antibodies to one don’t
protect against the rest,” said Ollmann Saphire. “Sudan virus is a particular
concern because it has caused about half of the ebolavirus outbreaks so far,
including the largest outbreak yet recorded.”

Uncovering the Body’s Natural Protection

US
government researchers recently demonstrated that an experimental vaccine
containing proteins from Ebola and Sudan viruses provides monkeys with some
protection against those viruses. But precisely how the vaccine works is
unclear, and it has never been tested in humans. Moreover, until now no
laboratory has isolated a neutralizing antibody against Sudan virus.

To
find such an antibody, Dye and his colleagues at Fort Detrick, Maryland,
injected lab mice with a harmless virus engineered to make copies of the Sudan
virus coat protein. The coat protein provoked the mice’s immune B cells to make
various antibodies against it, and the scientists were able to reproduce the
mice’s repertoire of antibodies by harvesting their B-cells and culturing them
in the lab. Testing each type of antibody for its ability to block the
infection of cells with Sudan virus, the researchers found one good candidate,
antibody 16F6, which not only neutralized Sudan virus in the lab dish but also
significantly delayed the deaths of infected mice. They then sent 16F6 to Ollmann
Saphire’s lab at Scripps Research in California.

"We
were very excited about developing this antibody as a potential treatment for
Ebola virus,” said Dye. “Collaborating with the Ollmann Saphire lab to
determine the binding site was the perfect complement to our previous
work"

Ollmann
Saphire’s lab specializes in the use of X-ray crystallography and related
techniques to visualize the atomic-scale details of viruses bound by
antibodies. These details reveal where on a virus an antibody binds, and if the
antibody is one that neutralizes a virus’s ability to infect cells, its binding
site usually offers important clues to the virus’s workings and
vulnerabilities.

In
the new study, Ollmann Saphire’s team found that 16F6 attaches to the Sudan
virus in a way that links two segments of the viral coat protein. The virus is
known to use one of these segments, GP1, to grab hold of a host cell. When this
happens, the cell automatically brings the virus inside, encapsulated within a
bubble-like chamber known as an endosome. Normally
the cell would destroy the contents of such an endosome, but Sudan virus–like
some other viruses–employs its other viral coat-protein segment, GP2, to fuse
to the wall of the endosome so that it and the rest of the virus escape into
the doomed cell’s interior. Antibody 16F6 seems to prevent this fusion process
from happening by keeping GP2 bound to GP1.

“The
virus is like a wolf in sheep’s clothing because its outer part is covered with
human sugar molecules, that the antibodies do not see as foreign,” said Ollmann
Saphire. “The binding site of the 16F6 antibody is one of the few places where
viral protein is exposed, and it’s exposed because it’s a place where GP1 and
GP2 need to be free to move.” To fuse to the endosomal wall, GP2 must separate
from GP1 and uncoil itself. When it is held fast to GP1 by the antibody 16F6,
GP2 can’t uncoil and perform its function–and so the Sudan virus, instead of
escaping into the relatively unprotected interior of the cell, stays within the
endosome and is eventually destroyed.

A Strategy Against Ebolaviruses

Ollmann
Saphire and her colleagues suspect that 16F6’s protein-linking strategy is the
best one that antibodies have against ebolaviruses. The antibody’s binding site
on the Sudan virus coat protein is virtually the same as the binding site of an
Ebola-Zaire-neutralizing antibody known as KZ52, which Ollmann Saphire and
Scripps Research colleague Professor Dennis Burton found and analyzed three
years ago. KZ52 is derived from antibodies made by an African patient who
survived an Ebola-Zaire outbreak in 1995, and aside from 16F6 it is the only ebolavirus-neutralizing
antibody whose binding site has been determined with X-ray crystallography.

“We
think it’s not just a coincidence that these two different antibodies, evoked in
two different host species by two different ebolaviruses, use the same strategy
of linking GP1 and GP2,” Ollmann Saphire said.

She
and her colleagues now are trying to obtain structural data on several other ebolavirus-neutralizing
antibodies, and she suspects that at least one of these also works by linking
GP1 to GP2. “There may be other neutralizing sites on ebolaviruses, but so far
the only one we’ve found is this one,” she said.

The
recognition that ebolavirus-neutralizing antibodies share this protein-linking
strategy should guide the further development of vaccines and immunotherapies.
“It helps us to understand more precisely what an ebolavirus vaccine or
immunotherapy ought to do,” Ollmann Saphire said.

The
lead authors of the paper, “A shared structural solution for neutralizing ebolaviruses,”
are João M. Dias, a research associate
in the Ollmann Saphire lab who is now a senior scientist at Heptares
Therapeutics in the UK; and Ana I. Kuehne, a researcher in the Dye laboratory
at Ft. Detrick. The other authors are Majidat A. Muhammad and Eugene Kang of
the Dye lab at Ft. Detrick; Dafna M. Abelson, Shridhar Bale and Marnie L. Fusco
of Scripps Research; Anthony C. Wong and Kartik Chandran of the Albert Einstein
College of Medicine; Peter Halfmann of the University of Wisconsin at Madison;
and Yoshihiro Kawaoka of the University of Tokyo. Funding for the research was
provided in part by the U.S. National Institutes of Health and the Defense
Threat Reduction Agency. For more information on the study, see http://www.nature.com/nsmb/journal/vaop/ncurrent/abs/nsmb.2150.html.

Send comments to: press@scripps.edu

The new Nature Structural & Molecular Biology study
shows how an antibody neutralizes the Sudan virus, a major species of
ebolavirus and one of the most dangerous human pathogens. (Image
courtsey of the Ollmann Saphire lab.)